Factor VII or VIIa—like molecules

Abstract

Conjugates of Factor VII (FVII) and Factor VIIa (FVIIA) are provided, as are methods for preparing them. Methods for producing novel polypeptides contributing to the production of such conjugates are provided. Methods of treatment by administering a FVII or FVIIa conjugate are provided.

1. A conjugate comprising:
 a polypeptide comprising an amino acid sequence which (a) differs from the hFVII or hFVIIa sequence SEQ ID NO:1 in 1-15 amino acid residues and (b) comprises an introduced in vivo N-glycosylation site relative to SEQ ID NO:1, wherein the introduced in vivo N-glycosylation site comprises the substitution K143N; and 
 a sugar moiety covalently attached to the introduced in vivo N-glycosylation site. 
 
     
     
       2. The conjugate of  claim 1 , further comprising at least one additional introduced in vivo N-glycosylation site. 
     
     
       3. The conjugate of  claim 2 , wherein the at least one additional introduced in vivo N-glycosylation site comprises the substitution I205S/T. 
     
     
       4. The conjugate of  claim 2 , wherein the at least one additional introduced in vivo N-glycosylation site comprises the substitutions R315N+V317S/T. 
     
     
       5. The conjugate of  claim 4 , wherein the polypeptide comprises the substitution V317T. 
     
     
       6. The conjugate of  claim 1 , wherein the polypeptide comprises the substitution N145S/T. 
     
     
       7. The conjugate of  claim 6 , wherein the polypeptide comprises the substitution N145T. 
     
     
       8. The conjugate of  claim 1 , wherein the polypeptide comprises the substitution K32D/E. 
     
     
       9. The conjugate of  claim 1 , wherein the polypeptide comprises the substitution A34D/E. 
     
     
       10. The conjugate of  claim 8 , wherein the polypeptide comprises an amino acid substitution at position 10. 
     
     
       11. The conjugate of  claim 2 , wherein the additional introduced in vivo N-glycosylation site comprises an amino acid substitution in a position selected from the group consisting of 28-48, 139-147, 286-294, 311-319, 338-345 and 388-406 relative to SEQ ID NO:1. 
     
     
       12. The conjugate of  claim 1 , further comprising at least one non-polypeptide moiety covalently attached to an amino acid residue of the polypeptide, wherein the non-polypeptide moiety is different from a sugar moiety. 
     
     
       13. The conjugate of  claim 12 , wherein the non-polypeptide moiety is a polymer molecule. 
     
     
       14. The conjugate of  claim 13 , wherein the polymer molecule is a linear polyethylene glycol or a branched polyethylene glycol. 
     
     
       15. The conjugate of  claim 14 , wherein the polyethylene glycol has a molecular weight of 300 Da to 100 kD. 
     
     
       16. A composition comprising the conjugate of  claim 1  and a pharmaceutically acceptable carrier or excipient. 
     
     
       17. A method for increasing blood clot formation in a mammal with a disease or condition in which increased clot formation is desirable, comprising administering to the mammal an amount of the conjugate of  claim 1  effective to increase blood clot formation. 
     
     
       18. The method of  claim 17 , wherein the disease or condition is selected from hemophilia, von Willebrand's disease, thrombocytopenia, severe tissue damage, trauma, and surgery.